ANAEROBIC CULTURE METHODS 163 



small inner tube, sterilized and inoculated. The pyrogallic 

 acid and sodium hydroxide are then placed in the bulb and 

 the stopper immediately replaced. 



This method has advantages over Buchner's in that the 

 oxygen is much more rapidly absorbed and the culture is 

 visible during incubation. 



Plates. 1. Ordinary deep culture (Petri) dish. 



2. McLeod's plate base (used with the bottom of a deep 

 Petri dish). (Muir and Ritchie, 6th Ed., Fig. 23, p. 66.) 



The principle of using these two plates is the same 

 throughout and is illustrated in Exercise 2. 



Jars. As has been noted before, the jars designed for 

 obtaining vacuum may be utilized in the pyrogallic acid 

 method and in the method making use of burning alcohol 

 to exhaust the oxygen. 



B. Liborius-Veillon Method and Roux's Biological 

 Method depend upon the abstraction of oxygen from the 

 medium by aerobic organisms. Liborius makes use of the 

 aerobes already present in the mixed culture, while Roux 

 uses a pure culture of an obligate aerobe. Nowak first grew 

 Bad. abortus by this method. 



Liborius-Veillon Method. 1. Fill long test tubes (22 

 cm. XI. 5 cm.) to a depth of 10-15 cm. with glucose agar or 

 gelatin and sterilize (below 120 C.). 



2. Place the tubes in a water bath and boil twenty to 

 thirty minutes to liquefy the agar and drive off the air dis- 

 solved in the medium; then cool to 40-45 C. until sown. 



3. Make loop dilutions in the melted agar and, as soon as 

 the tubes are sown, cool them rapidly in an upright position. 



Aerobic organisms grown in the upper part of the medium 

 which contains a certain amount of air in solution, while 

 the anaerobes multiply in the deeper layer. 



Roux's Method. 1. Make a deep agar or gelatin stab or 

 shake culture of the organism or substance to be studied. 



2. Pour upon the surface of this medium a layer 1 to 2 cm. 

 deep of a broth culture of a vigorous obligate aerobe as 



